Selenium Supplementation in Patients with Hashimoto Thyroiditis: A Systematic Review and Meta-Analysis of Randomized Clinical Trials

Background: Hashimoto thyroiditis (HT) is the most common cause of hypothyroidism in iodine-sufficient areas. Selenium is an essential trace element required for thyroid hormone synthesis and exerts antioxidant effects. Therefore, it may be of relevance in the management of HT. Methods: We conducted a systematic review and meta-analysis of randomized controlled trials (RCTs) to evaluate the effect of selenium supplementation on thyroid function (thyrotropin [TSH], free and total thyroxine [fT4, T4], free and total triiodothyronine [fT3, T3]), thyroid antibodies (thyroid peroxidase antibodies [TPOAb], thyroglobulin antibodies [TGAb], thyrotropin receptor antibody [TRAb]), ultrasound findings (echogenicity, thyroid volume), immune markers, patient-reported outcomes, and adverse events in HT. The study protocol was registered on PROSPERO (CRD42022308377). We systematically searched MEDLINE, Embase, CINHAL, Web of Science, Google Scholar, and the Cochrane CENTRAL Register of Trials from inception to January 2023 and searched citations of eligible studies. Two independent authors reviewed and coded the identified literature. The primary outcome was TSH in patients without thyroid hormone replacement therapy (THRT); the others were considered secondary outcomes. We synthesized the results as standardized mean differences (SMD) or odds ratio (OR), assessed risk of bias using the Cochrane RoB 2 tool, and rated the evidence using the Grading of Recommendations Assessment, Development, and Evaluation (GRADE) approach. Results: We screened 687 records and included 35 unique studies. Our meta-analysis found that selenium supplementation decreased TSH in patients without THRT (SMD −0.21 [confidence interval, CI −0.43 to −0.02]; 7 cohorts, 869 participants; I2 = 0%). In addition, TPOAb (SMD −0.96 [CI −1.36 to −0.56]; 29 cohorts; 2358 participants; I2 = 90%) and malondialdehyde (MDA; SMD −1.16 [CI −2.29 to −0.02]; 3 cohorts; 248 participants; I2 = 85%) decreased in patients with and without THRT. Adverse effects were comparable between the intervention and control groups (OR 0.89 [CI 0.46 to 1.75]; 16 cohorts; 1339 participants; I2 = 0%). No significant changes were observed in fT4, T4, fT3, T3, TGAb, thyroid volume, interleukin (IL)-2, and IL-10. Overall, certainty of evidence was moderate. Conclusions: In people with HT without THRT, selenium was effective and safe in lowering TSH, TPOAb, and MDA levels. Indications for lowering TPOAb were found independent of THRT.


Introduction
H ashimoto thyroiditis (HT), also referred to as chronic autoimmune or lymphocytic thyroiditis, is the most prevalent cause of hypothyroidism in iodine-sufficient areas. 1 It affects *160 million people globally, with women being 4-10 times more susceptible than men. 1,2HT is characterized by chronic inflammation of the thyroid gland, elevated serum antibodies against thyroid antigens, and typical appearance on thyroid ultrasound. 1 Once hypothyroidism develops, the current standard of care is lifelong thyroid hormone replacement therapy (THRT) with levothyroxine (LT4). 3ince several trace elements are essential for normal thyroid function, there is increasing interest in their supplementation for the management of HT, in particular the prevention of hypothyroidism. 4,5Besides iodine, one of the most discussed candidates is selenium. 6,7Selenium intake levels vary by region and are influenced by soil selenium content and selenium availability in the food chain, among other factors. 7Organ meats and seafoods are common sources of selenium, followed by muscle meats, cereals, and grains. 7he recommended daily allowance of selenium ranges between 55 and 70 lg for nonpregnant adults, which is often not reached, especially in Europe and some parts of China. 8,9any thyroid enzymes are selenoproteins, such as the deiodinases that metabolize thyroid hormones and the glutathione peroxidases (GPX) that help to manage oxidative stress in the thyrocyte. 7,10Reduced selenium levels have been observed in patients with autoimmune thyroid disease, including HT. 11 As a result, supplementing selenium in patients with HT has attracted much attention in recent decades.
So far, it has been suggested that selenium deficiency can exacerbate HT and the development of hypothyroidism. 2,12herefore, preventing selenium deficiencies could be a promising approach to prevent or modify HT-associated hypothyroidism.However, findings from previous systematic reviews and meta-analyses on the effect of selenium supplementation in HT remain inconclusive [13][14][15][16][17][18][19][20][21] due to factors such as a relatively small number of included studies, 13,[16][17][18][19] the inclusion of small and clinical heterogeneous populations (i.e., with and without THRT), 17 and outdated data due to the publication of new trials. 13,14,16,18,21In addition, the differences across studies in patient characteristics, outcome definitions, inclusion criteria, and supplementation regimens may result in considerable heterogeneity (I 2 ‡ 75%) and obscure the effect when results are statistically pooled. 22][25][26] In light of these inconclusive findings, we aimed to perform an updated systematic review and meta-analysis of randomized controlled trials (RCTs) investigating the effect of selenium supplementation on HT with particular attention to thyroid function, thyroid antibodies, ultrasound findings, immune markers, patient-reported outcome, and safety.We hypothesize that subgroup analyses will help us define indications for potential benefits.

Materials and Methods
We conducted this systematic review and meta-analysis in accordance with the current Preferred Reporting Items for Systematic Reviews and Meta-Analyses (PRISMA) 2020 guidelines. 27The processes and techniques complied with the Cochrane Handbook for Systematic Reviews of Intervention. 22We prospectively registered the study protocol on PROSPERO (CRD42022308377).A detailed description of the Supplementary Materials and Methods section is included in Supplementary Data S1.

Search strategy
We systematically searched the five electronic databases, MEDLINE, Embase, CINHAL, Web of Science, and Google Scholar, for publications and the CENTRAL Cochrane Library trial registry for ongoing and terminated trials from inception to April 5, 2022, and updated the search on November 14, 2023.We searched the Google Scholar database using the Publish or Perish software program 28 and limited reports to the first most relevant 200 on April 5, 2022.A medical information specialist designed the search strategy and developed the search strings (Appendix A1 and Supplementary Data S1).
We identified all citations (forward citation search) and references (backward citation search) of the included reports using the citation chaser app by Haddaway et al. and checked them for eligibility. 29

Study eligibility and data extraction
The following criteria qualified the studies for eligibility: (1) RCT, (2) selenium supplementation, (3) overt or subclinical hypothyroid or euthyroid, (4) with or without THRT, and (5) HT patients.There were no restrictions on publication date, publication language, participants' age, intervention duration, selenium regimen (type and dose), outcomes, and lack or presence of selenium deficiency.Outcomes included thyroid function (thyrotropin [TSH], free and total thyroxine [fT4, T4], free and total triiodothyronine [fT3, T3]), thyroid antibodies (thyroid peroxidase antibodies [TPOAb], thyroglobulin antibodies [TGAb], thyrotropin receptor antibody [TRAb]), ultrasound findings (echogenicity, thyroid volume), immune markers, patient-reported outcomes, and adverse events.TSH was a priori defined as a primary outcome due to its importance in evaluating disease progression; the others were considered secondary outcomes.When outcomes were thyroid function parameters, we performed separate main analyses in patients without THRT as markers of residual endogenous thyroid hormone production.
The two authors, V.V.H. and S.M.-A., independently screened titles and abstracts for eligibility and subsequently full texts if eligibility was not clear.Interrater disagreements were resolved through consensus or discussion with a third independent reviewer.We extracted the relevant data with a standardized, predesigned coding sheet (details in Supplementary Data S1).
We coded multiple intervention groups from a single study separately, hereafter referred to as cohort, and combined cohorts that had a common control group to create single pairwise comparisons. 22Missing data were obtained directly from the authors.When the authors could not provide the missing results, we extracted the results from the diagrams using Rohatgi's WebPlotDigitizer, available at https://auto meris.io/WebPlotDigitizer(2022), where possible.

Data synthesis and statistical analyses
We performed a meta-analysis when at least two cohorts provided poolable results for an outcome, and a subgroup analysis and meta-regression when more than 10 cohorts were available. 22We conducted all the statistical analyses in R version 4.1.2(R Core Team) using the metafor package. 30e statistically analyzed results as standardized mean differences (SMD) using the random-effects model, accounting for systematic variations in effect sizes between studies. 31,32For dichotomous outcomes, we analyzed odds ratios (ORs).Publication bias was assessed using funnel plot asymmetry, 33 the Egger's regression, 34 and rank correlation tests 35 (details in Supplementary Data S1).
To identify sources of between-study heterogeneity, we performed stratified analyses, evaluating the potential impact of the following: Selenium dose (elemental selenium) Intervention duration Thyroid status (overt hypothyroidism, subclinical hypothyroidism, euthyroidism) Sex distribution (percentage of females) Participant age group ( ‡18 years, <18 years, mixed population) THRT Selenium status (severely selenium deficient, mildly selenium deficient, and selenium sufficient) Blinding Risk of publication bias.
In addition, we performed random-effects meta-regression analyses with selenium dose, intervention duration, sex distribution, and serum selenium levels as independent variables and effect estimates as dependent variables.p-Values below 0.05 were considered significant. 30To evaluate the impact of individual studies on the overall results, we visually inspected the Baujat plot 36 and assessed the pooled risk estimates and heterogeneity after removing studies from the analyses one by one from (leave-one-out analysis) using the influence function. 37

Quality assessment
Two reviewers (V.V.H. and S.M.-A.) individually assessed the risk of bias in the included studies using the Cochrane Revised Risk of Bias Tool for randomized clinical trials (RoB 2). 38

GRADEing of evidence
The two authors, V.V.H. and S.M.-A., assessed the certainty of the evidence for each outcome in our meta-analysis for the longest time point using the Grading of Recommendations Assessment, Development, and Evaluation (GRADE) method. 39

Study selection
We identified 1025 records, of which 687 remained after deduplication (312 duplicates).We checked the titles and abstracts, and if necessary the full texts, and identified 60 reports.In the full-text screening, 26 reports were exclu-ded (Supplementary Table S1 in Supplementary Data S2).We identified one additional record by Shabalina and Fadeyev during the citation searching, which was not present in the databases searched. 40No additional study was included after cross-checking studies from previous systematic reviews and meta-analyses.A trial by Bonnema and colleagues was identified from the Cochrane Library Central, 41 which confirmed that the recruitment was complete and publication was expected at the end of 2023.However, preliminary data were not available for this meta-analysis.Finally, we included 35 unique studies in the systematic review and 32 in the meta-analysis (Fig. 1).
Of the 35 studies, 30 (86%) diagnosed HT using TPOAb presence.The diagnostic thresholds for TPOAb levels varied (Supplementary Table S2 in Supplementary Data S2).Euthyroidism, subclinical hypothyroidism, and overt hypothyroidism were consistently defined based on TSH and fT4 levels, with euthyroidism defined as normal TSH levels and normal fT4 levels, subclinical hypothyroidism defined as elevated TSH levels with normal fT4 levels, and hypothyroidism defined as elevated TSH levels with decreased fT4 levels.However, the reference ranges for fT4 and fT3 levels varied across studies.The TSH reference ranges were predominantely consistent, mostly below or equal to 4 mIU/L, with a few studies extending the range up to 5 mIU/L, which could be due to assay differences.Eight studies (23%) did not report the TSH reference ranges (Supplementary Table S3 in Supplementary Data S2).

Effect of selenium supplementation on thyroid function
A total of 32 cohorts investigated the effect of selenium supplementation on at least one thyroid parameter (TSH, fT4, fT3, T4, T3) (Table 1).
Thyrotropin.From the 11 cohorts reporting on TSH levels in participants without THRT, one observed a decrease 70 and ten observed no change [43][44][45][46]58,61,65 in TSH levels after SELENIUM IN HASHIMOTO THYROIDITIS selenium supplementation compared with the control group. Among th 20 cohorts reporting on TSH levels in participants with THRT or unspecified THRT, 4 observed a decrease, 40,52,62,64 1 an increase, 66 and 15 no change in TSH levels after selenium supplementation compared with the control group.The meta-analysis demonstrated a significant decrease in TSH levels after selenium supplementation compared with the control group in patients without THRT (SMD -0.21 [confidence interval, CI -0.41 to -0.02]; 7 cohorts; 869 participants; I 2 = 0%) (Table 2 and Fig. 2).
The TSH levels in the overall population remained unchanged (SMD -0.S5 and Supplementary Fig. S18 in Supplementary Data S2).The selenium compound significantly affected the results ( p = 0.03).No outlier cohort was identified (Supplementary Fig. S23 in Supplementary Data S2).Nevertheless, due to missing data in the original publication and conflicting result estimates, we excluded the study by Esposito et al. 58 We rated quality of evidence as moderate (Supplementary Table S6 in Supplementary Data S2).
Free thyroxine.From the 23 cohorts that reported fT4 levels, one observed an increase, 46 one a decrease, 40 and 21 no change in fT4 levels after selenium supplementation.The meta-analysis showed no significant increase in fT4 levels FIG. 1. Flowchart for inclusion of the 35 studies adapted from the PRISMA 2020 statement. 27Six databases were systematically searched during the identification phase.Titles and/or abstracts were screened, and if records were eligible, the full text was assessed during the screening phase, resulting in the total number of studies included.Records refer to the title and/or abstract of a report.A report is a document that provides information about a study.AE, adverse event; Echo, ultrasound echogenicity; fT3, free triiodothyronine; fT4, free thyroxine; IM, immune markers including glutathione peroxidase, malondialdehyde, superoxide dismutase, total antioxidant capacity; PRISMA, Preferred Reporting Items for Systematic Reviews and Meta-Analyses; QoL, quality of life; T3, total triiodothyronine; T4, total thyroxine; TGAb, thyroglobulin antibodies; TPOAb, thyroid peroxidase antibodies; TRAb, thyrotropin receptor antibody; TSH, thyrotropin; Vol, thyroid volume.Removing the two outlier cohorts 46,61 led to similar results (Supplementary Fig. S23 in Supplementary Data S2).We rated quality of evidence as moderate (Supplementary Table S7 in Supplementary Data S2).
Free triiodothyronine.fT3 levels increased significantly in 2 out of 11 cohorts (18%) after intervention, 53,58 while no significant effect was observed in the remaining nine cohorts (82%).The meta-analysis showed no significant increase in fT3 levels after selenium supplementation compared with the control group (SMD 0.51 [CI -0.11 to 1.13]; 11 cohorts; 658 participants; I 2 = 84%) (Table 2 and Supplementary Fig. S6 in Supplementary Data S2), with similar results when restricting the analyses to cohorts without THRT (Table 2 and Supplementary Figs.S7 and S8 in Supplementary Data S2).Subgroup analysis did not reveal any significant influencing factors on the outcome (Supplementary Table S5 and Supplementary Fig. S22 in Supplementary Data S2).Removing the outlier cohort 58 returned comparable results (Supplementary Fig. S23).We rated quality of evidence as very low (Supplementary Table S6 in Supplementary Data S2).
Total thyroxine and total triiodothyronine.T4 was reported in three 67,71 and T3 in four cohorts, 56,67,71 showing no significant difference between the selenium-treated and control groups (Supplementary Figs.S9 and S10 in Supplementary Data S2).

Effect of selenium supplementation on thyroid antibodies
Our systematic review included 34 cohorts that investigated the impact of selenium supplementation on thyroid antibodies such as TPOAb, TGAb, and TRAb.
Removing the outlier cohort 64 returned comparable results (Supplementary Fig. S23 in Supplementary Data S2).We rated quality of evidence as low (Supplementary Table S6 in Supplementary Data S2).
In pregnant women, TPOAb titers decreased significantly at delivery 51 and 5 or 6 months postpartum, 50,51 but no significant effect was observed during gestation 49,50 (Supplementary Fig. S11 in Supplementary Data S2).
Thyrotropin receptor antibody.One study assessed TRAb and found a significant decrease in the seleniumtreated compared with the control group. 53fect of selenium supplementation on ultrasound findings Thyroid echogenicity.Eight cohorts investigated the effect of selenium on ultrasound echogenicity.Thyroid echogenicity decreased significantly after selenium supplementation (25% and 50% of participants) compared with the control group (5% and 12% of participants) in two cohorts (33%) 57,59 with no effect in the other six (67%). 40,45,50,58yroid volume.Seven cohorts evaluating the effect of selenium on thyroid volume did not find significant results. 40,45,48,50,64The meta-analysis of the poolable cohorts indicated the same effect (SMD -0.14 [CI -0.57 to 0.28]; 4 cohorts; 182 participants; I 2 = 0%) (Supplementary Fig. S13 in Supplementary Data S2).

Effect of selenium supplementation on immune markers
In total, 18 cohorts assessed the inflammatory and antioxidant markers such as interleukins (ILs), chemokines, cytokines, and selenoproteins, with mixed results across studies.

Effect of selenium supplementation on patient-reported outcomes
A total of six cohorts evaluated the effect of selenium supplementation on well-being in HT patients. 45,50,54,57,65Of these, two cohorts measuring well-being with either an unspecified questionnaire or the short form 12 health survey (SF-12) questionnaire reported a higher percentage of improvement in well-being in selenium-treated patients compared with controls. 54,57The other four cohorts, measuring well-being with the short form 36 health survey questionnaire (n = 3) 45,65 or the SF-12 questionnaire (n = 1), 50 found no significant difference in quality of life between the two cohorts.All six cohorts were blinded.

Safety of selenium supplementation
2]74 Of these, 13 cohorts reported no adverse events.Five cohorts listed between 2 and 19 adverse events, including nausea, vomiting, fever, dizziness, chest tightness, bloating, gastrointestinal problems, hair loss, and miscarriages. 43,51,65,71one of the studies assessed events for severity and causality as recommended by the ICH E2A guidelines. 77Our metaanalysis found no significant difference in adverse events between the selenium and control groups in the studies with adverse events (OR 0.89 [CI 0.46 to 1.75]; 16 cohorts; 1339 participants; I 2 = 0%) (Supplementary Fig. S17 in Supplementary Data S2).We rated quality of evidence as moderate (Supplementary Table S6 in Supplementary Data S2).

Risk-of-bias assessment
Of the 31 studies reporting on TSH (i.e., our primary outcome), 12 (39%) raised some concerns, while 19 (61%) had a high risk of bias (Supplementary Table S7 in Supplementary Data S2).Most of these concerns originated from the randomization process and the selection of the reported results.9][70] Eight (26%) of the studies reported the details of the randomization process. 40,48,58,61,65,70,71,76Other common issues included a lack of power analysis and a clear description of the statistical analysis.Sixteen (52%) measured the serum selenium concentration and found a significant increase in seleniumtreated participants to around 100 lg/L, while the control group showed no changes.Only one study found no significant increase in the selenium-treated group 67 (Table 1).The overall certainty of evidence was moderate (Supplementary Table S6 in Supplementary Data S2).

Assessment of publication bias
Supplementary Figure S24 in Supplementary Data S2 depicts the funnel plots for detection of publication bias.Egger's regression tests and the rank correlation test did not show statistically significant funnel plot asymmetry for any outcomes, except for TPOAb, where the rank correlation indicated significant funnel plot asymmetry ( p = 0.04) (Table 2 and Supplementary Fig. S24 in Supplementary Data S2).Results remained similar after restricting the analyses to cohorts within the funnel ( p > 0.05) (Supplementary Fig. S24 in Supplementary Data S2).

Discussion
This systematic review and meta-analysis showed a significant reduction in TSH levels following selenium supplementation in HT patients without THRT, and these effects disappeared when including patients receiving THRT.Furthermore, selenium supplementation exhibited favorable results by reducing TPOAb and MDA levels, with no statistically significant effect on fT4, fT3, T4, T3, TGAb, thyroid volume, and IL-2 and IL-10.Selenium supplementation was well tolerated, as evidenced by the absence of significant differences in adverse events between the selenium and placebo groups.Overall, the certainty of evidence was moderate.
Previously, three meta-analyses have investigated the effect of selenium on TSH levels in autoimmune thyroiditis patients. 13,15,21One of these meta-analyses included exclusively LT4-untreated patients, 13 one included LT4treated and LT4-untreated patients, 21 and the other additionally included patients with Graves' disease and hyperthyroidism. 15Neither meta-analyses found a significant effect of selenium on TSH levels.However, two of the prior meta-analyses were constrained by statistical power, including only five and eight trials, respectively. 13,15Furthermore, they exhibited considerable and unexplained heterogeneity (I 2 = 94%, 76%, and 55%, respectively) and lacked subgroup analyses.Against this background, our meta-analysis adds to the existing knowledge in this field by demonstrating an effect of selenium supplementation on lowering TSH levels exclusively in HT patients without THRT.Moreover, the quality of evidence assessed in our meta-analysis was moderate, which is an improvement from the quality of evidence previously assessed (i.e., low to very low). 1319]21 Of these, two meta-analyses consistently reported significant effects at 3, 6, and 12 months, 14,21 two meta-analyses detected a significant effect solely at 6 months, not at 3 months, 17,19 and the remaining two meta-analyses did not differentiate between time points. 15,18Against this background, our meta-analysis showed a consistent reduction in TPOAb levels, with significant reductions observed at 3-4 months and after at least 6 months of selenium supplementation.8][19] This expanded data set enabled us to address the substantial heterogeneity observed in both our analyses (I 2 = 90%) and earlier meta-analyses (I 2 = 99%, 15 94%, 19 67-95%, 17 23-97%, 14 63%, 18 95% 21 ) through extensive subgroup and meta-regression analyses.
Our study provides a notable advance over previous systematic reviews and meta-analyses that were limited in scope and focused primarily on a restricted set of outcomes, such as thyroid function parameters (TSH, fT4, fT3), 13,15 thyroid antibodies (TPOAb, TGAb), 14,15,17,19 and, in two cases, mood, with limited cohort inclusion. 17,18We expanded the scope to encompass a broader range of thyroid function parameters, including T4 and T3, and ultrasound findings such as echogenicity and thyroid volume.We also included immune markers, revealing a significant reduction in MDA levels, a marker of oxidative stress. 78Importantly, our evidence indicated the safety of selenium supplementation at doses ranging from 80 to 400 lg per day for up to 12 months by showing comparable adverse events between the selenium-treated and control groups, filling a notable gap where such safety analyses were lacking.Adverse effects associated with adequate selenium supplementation are rare in the literature.
However, excess intake of selenium supplementation can be detrimental, with symptoms of acute toxicity (e.g., gastrointestinal and neurological manifestations) typically occurring at doses of 300-400 lg/day. 8,9The European Food Safety Authority (EFSA) has set the tolerable upper intake level for selenium at 255 lg/day. 79One clinical trial raised concerns that prolonged selenium supplementation of 200 lg/day could increase the incidence of type 2 diabetes. 80owever, it should be noted that the overall study population was not initially selenium deficient, the supplementation duration was extended (7.7 years), and diabetes was a secondary outcome, making interpretation difficult.Furthermore, it has been recommended that people with serum or plasma selenium concentration of at least 122 lg/L should not supplement selenium as this may increase the risk of adverse events, including cancer and type 2 diabetes. 7,81ur detailed subgroup analyses for TSH, fT4, TPOAb, and TGAb identified patient groups and intervention types associated with the most robust results and explored factors contributing to heterogeneity.When we restricted our analysis to adult participants ( ‡18 years), selenium supplementation led to a reduction in TSH and TPOAb levels and an increase in fT4 levels.In contrast, results remained insignificant in minors (<18 years).Notably, only four of the included cohorts investigated minors, suggesting that the lack of a significant effect may be due to insufficient statistical power.We observed consistently stronger or similar results after limiting the analysis to cohorts with the selenium compound selenomethionine.Selenomethionine, an organic form of selenium, is the most commonly used form for dietary supplementation.Although inorganic selenium may have a greater ability inserted into GPX, organic forms dominate in their capability to be stored and integrated into body proteins. 82ubgroup analysis based on initial selenium status did not reveal clear trends.A possible explanation may be that data on selenium-sufficient cohorts were scarce.Most studies were conducted in populations most at risk for selenium deficiency, that is, the European and Chinese women. 12,83nly half of the studies reported selenium levels at baseline, of which 89% of the cohorts were selenium deficient.However, based on epidemiological data from China, Wu et al. postulated that selenium deficiency is a modifiable risk factor for HT. 12 As deficiencies are expected to increase further with the rising popularity of plant-based diets, particularly among women, 84,85 the recommended daily intake of *55-70 lg/day should be ensured, with higher intakes required during pregnancy and lactation. 9The female preponderance in our systematic review may be partly explained by the increased incidence and prevalence of HT in women.
However, the underrepresentation of male patients may lead to gender disparities in the management of HT.None of the studies analyzed the data by sex, highlighting the need for further investigation into potential sex differences.As exposure to sex hormones may influence autoimmunity, future investigations must also account for menopausal status (including the use of THRT). 86The high heterogeneity of results on TPOAb and TSH levels described in previous and in our current meta-analysis [13][14][15]17,19 was reduced when we restricted our results to cohorts with severe selenium deficiency and to female cohorts only. This ighlights the importance of adjusting for sex and selenium status of participants in future research.
Notably, no significant effects on TSH were observed when including participants receiving THRT.TSH levels may be difficult to interpret during LT4 substitution, as they primarily depend on replacement doses, and none of the included studies described a correction or washout procedure before laboratory quantification.In contrast, the effect on TPOAb was more pronounced in our study when we focused on HT patients initially treated with LT4 who had hypothyroidism.However, LT4 treatment may not necessarily reflect a decrease in the autoimmune response, but rather a normalization of thyroid function. 87Addressing the issue of LT4 dose adjustment may enhance the robustness of future trials.No significant increases in the thyroid hormones fT4 and fT3 were observed.This could be due to the fact that participants without THRT were mostly euthyroid or subclinically hypothyroid, that is, fT4 and fT3 levels were in the normal range (*2-7 and 10-24 pmol/L, respectively).
HT is a complex autoimmune disease with pathophysiological mechanisms involving a feedback loop of immune system cells and cytokines.T cells are activated, leading to the production of inflammatory cytokines (e.g., IFN-c and TNF-a) and the release of chemokines (e.g., CXCL-10) by thyrocytes, which further amplify the inflammatory response and attract additional T cells.This promotes the production of antibodies against thyroid-specific proteins (e.g., TPOAb, TGAb, TRAb), thyroid damage, and further development of hypothyroidism. 1,2,6In patients with HT, selenium supplementation can reduce TPOAb and TSH concentrations through several mechanisms, which can be related to the antioxidative and anti-inflammatory role of selenoproteins.Most selenoproteins are expressed in the thyroid gland, including the iodothyronine deiodinases, GPX, and selenoprotein S.These proteins perform essential functions such as thyroid hormone metabolism and activation, protection against oxidative damage, and regulation of the inflammatory response. 11,88elenoenzymes, such as GPX, play an antioxidative role by reducing the formation of free radicals and hydrogen peroxides. 46Selenoenzymes also play a beneficial role in the immunoregulatory process, involving T cell activity and cytokine production. 70In line, several studies included in our systematic review showed that selenium supplementation increased GPX activity, 46,70,76 decreased inflammatory and oxidative activity (IL-2, MDA, IFN-c, and TNF-a), 45,46,52,71,72,74 and increased antioxidant activity (IL-10, SOD, and TAC). 71,72Of note, there may be interindividual differences in the extent to which TPOAb concentrations change in response to selenium supplementation. 46olymorphisms within selenoprotein genes impact not only SELENIUM IN HASHIMOTO THYROIDITIS their structure and function, but also individual responses to selenium intake.Consequently, some individuals may exhibit modulated inflammatory responses, 89 endoplasmic reticulum stress in the brain, 90 susceptibility to selenium deficiency, and specific response to selenium supplementation. 46his review has identified several limitations of the available evidence on the effects of selenium supplementation on HT.While measurements of TSH, fT4, and fT3 are more harmonized, the included studies used different assays to assess TPOAb and TGAb to measure distinct subgroups of antibodies with varying affinities. 91To address this issue, we synthesized the results of the studies using the SMD instead of the mean difference, which is recommended when varying scales and assays were used in the included studies.However, only the effect size can be interpreted and not its clinical significance. 22In addition, serum selenium concentrations can vary substantially depending on the analytical technique.Selenium status is typically assessed through the quantification of extracellular selenium levels in plasma or serum, reflecting the overall selenium intake within the past several days. 7,92n contrast to selenium in blood, intracellular selenium levels in erythrocytes, measured via inductively coupled plasma mass spectrometry, remain unaffected by the inflammatory response.This implies potential advantages, but standardized assessment is lacking. 92The general lack of harmonization may be a major cause of the large heterogeneity in the meta-analyses.
Another limitation of the studies included in this systematic review is the lack of information on the dietary habits of the participants.Selenium is present in various animal and plant food sources, and other dietary components, such as iodine or iron, can affect thyroid function and autoimmunity. 7,93Future studies should therefore report and account for baseline selenium, iodine, and iron status when analyzing the results.The same applies to the compliance, which was often not reported.Methods such as pill counts or evaluations of serum selenium levels should be used to assess compliance.In addition, the dose and regimen of THRT were not considered, which could significantly affect thyroid hormone levels, especially if THRT was taken immediately before the study assessments.
Finally, all included studies raised at least some concerns about the risk of bias according to the RoB 2 assessment.The methodology, for example, the randomization process and the selection of reported outcomes, was mostly not explained in detail, which should be improved in further studies.However, the leave-one-out analysis identified very few outlier cohorts, and exclusion produced similar results, leading to robust findings that were confirmed by the GRADE assessment.

Conclusion and Implications for Practice, Policy, and Future Research
Our systematic review and meta-analysis provide valuable insights for clinical practice, policy, and future research.
Clinically, selenium supplementation showed promise in reducing TSH levels, especially in euthyroid and subclinically hypothyroid individuals without THRT.Moreover, selenium supplementation appears to have a beneficial effect in lowering TPOAb levels, although the clinical relevance of lowering TPOAb levels warrants further investigation. 94elenium doses above 100 lg/day may be most potent.
In terms of policy implications, standardizing quantification techniques for thyroid hormones and antibodies is crucial for accurate and comparable measurements.
We recommend that future studies include selenium level assessment during the study, stratify data by sex, selenium status, and thyroid status, and provide clear reference values for all parameters.High-quality studies with larger sample sizes and detailed reporting according to Consolidated Standards of Reporting Trials guidelines 95 are essential to confirm and fully understand selenium's role in HT.Research should also include children, adolescents, and pregnant women, and investigate the long-term effects of selenium supplementation on hypothyroidism development from euthyroidism.
In conclusion, our study suggests that selenium supplementation is safe and holds potential as a disease-modifying factor for HT-associated hypothyroidism.Further research is needed to confirm its efficacy, fully understand its mechanism of action, and elucidate its cost-effectiveness.

FIG. 3 .
FIG. 3. Effect of selenium supplementation on TPOAb (IU/mL), stratified by thyroid hormone replacement therapy, in Hashimoto thyroiditis (n = 2358).Black rectangles represent SMD for each study; the size of the rectangle is proportional to the weight of the study for the pooled effect.Horizontal lines indicate CI.The black diamond summarizes the pooled SMD data.(1)/(2) Indicate cohorts 1 and 2 of the study.

Table 1 .
Study and Participant Characteristics of the 35 Included Randomized Controlled Trials
a Only participants receiving same intervention throughout the entire study considered.b Results not available.c Data estimated from other studies.d Results extracted from graphs.e Participants already on THRT at study start and therefore reported euthyroid at study start.f One intervention group and its corresponding comparison group additionally received 30 mg zinc per day.g Time point 12 months after delivery was not included in our analysis to increase comparability with other studies.
-1.16 [-2.29 to -0.03] 85 ---Bold values indicate significant results.CI, confidence interval; OR, odds ratio; SMD, standardized mean difference.FIG. 2. Effect of selenium supplementation on TSH (mIU/L), stratified by thyroid status, in patients with Hashimoto thyroiditis without thyroid hormone replacement therapy (n = 869).Black rectangles represent SMD for each study; the size of the rectangle is proportional to the weight of the study for the pooled effect.Horizontal lines indicate CI.The black diamond summarizes the pooled SMD data.CI, confidence interval; Control, Control group receiving placebo or nothing; SMD, standardized mean differences.after